Search Results (3,305)

One highly consequential presentation of Venezuelan equine encephalitis virus (VEEV) infection is encephalitis. Here we considered anti-inflammatory interventions to limit the effects of this using a BALB/c subcutaneously challenged mouse model of disease. This disease model nearly ubiquitously presents with severe encephalitis, where viral neuroinvasion correlates with much of the outward clinical signs of disease. A selection of already licenced, commonly used anti-inflammatory drugs were tested in mice developing encephalitis (starting treatment at post challenge). Drug regimens were used that had previously been shown to have pharmacodynamic effects in mice for unrelated conditions. None of the treatment regimens tested reduced brain inflammation. A single anti-inflammatory drug (dexamethasone) was further tested utilising ascending doses in an effort to provide an effective anti-inflammatory regimen. Higher doses of dexamethasone ( and ) reduced inflammatory markers in the brain and lowered weight loss and clinical signs early on during infection. However, the regimen also caused the disease to become more severe at later time points when compared to controls. When combined with the antiviral drug molnupiravir, the negative effects of the dexamethasone treatment ( and ) were absent, and the positive disease severity-reducing effects remained. When combined with a specific VEEV monoclonal antibody (1A3B7), dexamethasone significantly reduced the antibody’s protective effects. These data present currently unique insights into how anti-inflammatory approaches might benefit patients with VEEV disease and where caution might be advised. Full article

Background/Objective: The influenza virus remains one of the most prevalent respiratory pathogens, posing significant global health and economic challenges. According to the World Health Organization, the seasonal influenza virus infects up to 1 billion people and causes up to 650,000 deaths, annually. Despite influenza vaccination is the most effective available preventive strategy, its reliance on strain predictions and yearly updates limits its effectiveness. The virus’ ability to cause both epidemics and pandemics, driven by zoonotic transmissions, underscores its continuous threat. The ongoing H5N1 avian influenza outbreak is the perfect example, renewing concerns due to its ability to infect over 70 mammalian species and sporadically transmit to humans. This study aims to evaluate the protective potential of two human monoclonal antibodies against diverse and recent influenza virus strains. Method: PN-SIA28 and PN-SIA49 monoclonal antibodies were previously isolated from an individual undergoing seasonal influenza vaccination and with no known recent influenza virus exposure. Their breadth of recognition, neutralization, and conferred in vivo protection were assessed against multiple influenza viruses, including pre-pandemic strains. Structural analyses were performed to characterize antibody–antigen interactions for epitope identification. Results: Both antibodies recognize a broad range of strains and neutralize pre-pandemic avian influenza viruses, including the currently circulating H5N1 clade. Moreover, a structural analysis revealed that PN-SIA49 binds a conserved HA stem region, overlapping with epitopes recognized by other broadly neutralizing antibodies. Conclusions: These findings underscore the potential of broadly neutralizing antibodies as a basis for universal influenza countermeasures against both seasonal and pandemic threats. Additionally, they provide guidance for the design of targeted vaccine strategies to steer immune responses toward broadly protective epitopes. Full article

Background: Non-typhoidal Salmonella (NTS) is a major foodborne pathogen, with poultry products, especially eggs, being the primary source of human infections. Current serovar-specific poultry vaccines effectively reduce targeted Salmonella serovars but may inadvertently promote the emergence of untargeted serovars within poultry flocks. Therefore, novel vaccine candidates providing broad cross-serovar protection are needed to improve overall effectiveness of Salmonella control programs. Objectives: This study evaluated the immunogenicity of the novel subunit vaccine candidate InvG and assessed its ability to reduce Salmonella colonization in vaccinated laying hens and their progeny through maternally derived antibodies transferred via egg yolk. Methodology: Three experiments were performed. Experiment I evaluated the immunogenicity of purified recombinant InvG by (a) measuring anti-InvG antibodies using an enzyme-linked immunosorbent assay (ELISA) and (b) completing transcriptomic profiling of immune responses in vaccinated chickens. Vaccinated chickens were subsequently challenged with Salmonella Enteritidis to assess the efficacy of anti InvG antibodies in reducing intestinal colonization of Salmonella. Experiment II involved immunizing hens with InvG, to evaluate passive transfer of antibodies via egg yolk and the protective efficacy of maternally derived antibodies against Salmonella challenge. Passive transfer was assessed by measuring IgY antibodies in hen serum, egg yolk, and progeny serum, as well as secretory IgA (sIgA) antibodies in progeny intestinal washings using ELISA. Protective efficacy was evaluated by orally challenging one-day-old chicks with three different Salmonella serovars. Experiment III assessed the persistence of anti-InvG antibodies in the serum of vaccinated hens and their transfer into eggs following two doses of InvG. Results: InvG vaccination induced robust IgY antibody responses in hens, with efficient maternal antibody transfer to progeny via egg yolk. A statistically significant reduction in Salmonella colonization was observed in both vaccinated hens and their progeny. Conclusions: These findings demonstrate that InvG represents a promising subunit vaccine candidate for Salmonella control in poultry and warrants further investigation towards development as a broadly protective commercial poultry vaccine against Salmonella. Full article

Background: Emerging immune-evasive viral variants threaten the efficacy of current vaccines, underscoring the need for strategies that elicit broad and durable protection. Heterologous prime–boost regimens combining distinct vaccine platforms can enhance humoral and cellular immunity through complementary mechanisms. Methods: Using an intramuscular immunization scheme aligned with clinical vaccination practice, CD-1 mice received homologous or heterologous prime–boost regimens combining a replication-deficient Orf virus (Parapoxvirus orf, ORFV)-based spike vaccine (ORFV-S) with the licensed adjuvanted recombinant protein vaccine VidPrevtyn Beta. Spike-specific humoral and cellular immune responses were assessed. Results: ORFV-S alone induced potent and broad spike-specific IgG responses and achieved the strongest ACE2-binding inhibition across variants of concern. ORFV-S priming followed by VidPrevtyn Beta boosting markedly enhanced the magnitude and cross-variant breadth of antibody responses compared with homologous protein vaccination. Both homologous ORFV-S and heterologous regimens incorporating ORFV-S elicited strong CD4⁺ and CD8⁺ T-cell responses, whereas VidPrevtyn Beta alone induced only modest T-cell activity, demonstrating that ORFV-S effectively complements protein-based vaccines. Conclusions: The ORFV-S vector represents a potent vaccine platform capable of inducing broad humoral and cellular immunity. Its use in heterologous prime–boost combinations enhances both antibody magnitude and breadth beyond homologous protein vaccination, supporting its application in vaccination strategies against evolving viral pathogens. Full article

Background: Patients with end-stage renal disease (ESRD) on hemodialysis are at increased risk for severe influenza, and underlying immune dysfunction may limit vaccine-induced protection. Methods: This observational open-label study evaluated immune responses in 93 hemodialysis patients vaccinated with seasonal inactivated influenza vaccine (IIV) during the 2019–2020 (n = 22) and 2023–2024 (n = 71) seasons. Immune responses were comprehensively assessed using hemagglutination inhibition and microneutralization assays to measure antibody levels, together with flow cytometry analysis of key immune cell populations, including plasmablasts, T-follicular helper cells (Tfh), and effector memory T cells (Tem). Results: During the 2019–2020 season, antibody responses in hemodialysis patients were comparable to those in healthy volunteers in both younger (18–60 years) and older (over 60) age groups. By day 7 post-vaccination, there was a pronounced increase in activated Tfh1 cells, coinciding with a surge in plasmablasts and a rise in antigen-specific B cells. This was accompanied by a T-cell response mediated by IFNγ-producing and polyfunctional CD4+ Tem cells. In the 2023–2024 season, revaccination was associated with higher baseline antibody levels but did not alter subsequent response kinetics to A/H1N1pdm, A/H3N2, and B/Yamagata antigens. In contrast, responses to B/Victoria were higher in revaccinated patients throughout the entire observation period. Conclusions: Our findings confirm that standard-dose IIV vaccination is beneficial for hemodialysis patients, inducing robust and adequate humoral and T-cell immune responses. Full article

Background: Studies comparing systemic and salivary antibody responses against SARS-CoV-2 between children and adults show conflicting results. Furthermore, it is still unclear whether salivary antibody testing could be a non-invasive approach to evaluate the humoral immune response. Methods: anti-SARS-CoV-2 IgG antibodies were measured in blood and saliva sample pairs from vaccinated adults to investigate whether salivary antibody response could be a non-invasive assessment of immune response. Salivary antibody levels were also compared between vaccinated children and adults to investigate local antibody responses. Results: Salivary IgG antibody response against SARS-CoV-2 largely reflects the systemic response in vaccinated adults. Salivary and systemic antibody concentrations were higher in vaccinated adults who had been infected, received schemes including mRNA-based vaccines, had more exposures, and a shorter time from last exposure. Salivary antibody detection was associated with schemes including mRNA-based vaccines, time from last exposure, and systemic antibody concentrations. Vaccinated children showed higher salivary antibody concentrations than adults. This difference remained when comparing antibody levels between children and adults under equal conditions (vaccination schemes, number of exposures, time from last exposure, COVID-19 history). Younger age, number of exposures, schemes including mRNA-based vaccines, and shorter time from last exposure were associated with salivary antibody levels in a multivariable linear regression analysis (p < 0.0001). Conclusions: Salivary antibody determination against SARS-CoV-2 could be a non-invasive assessment of the short-term immune response in adults with multiple exposures. Furthermore, the stronger salivary antibody response in children suggests that local immune protection may differ between children and adults, contributing to different outcomes. Full article

Background: Tumor-lysate vaccines can capture tumor heterogeneity; however, their effectiveness may be reduced by antigen instability and short antigen presentation. Here, we aimed to improve antigen protection and prolong presentation by using a slow-degrading polymeric nanocarrier and an approved adjuvant. Methods: We encapsulated breast cancer cell lysates (MCF-7 and MDA-MB-231) in poly(ε-caprolactone) (PCL) nanoparticles using a double-emulsion (w/o/w) method and co-administered them with alum. We then characterized particle size, PDI, zeta potential, morphology, and in vitro release. Next, we evaluated nitric oxide (NO), TNF-α/IL-10 responses, and cytocompatibility in J774 macrophages. Finally, we quantified serum antibody titers in Balb/c mice after six biweekly immunizations, generated hybridomas, purified IgG, and tested antibody-mediated cytotoxicity alone and together with doxorubicin. Results: PCL nanoparticles were ~220–255 nm (PDI 0.10–0.19; ζ −2 to −3 mV) and released ~90–95% of encapsulated lysate by 800 h (~33 days). Encapsulated lysate (40 μg/mL) modestly increased NO versus control and increased further with alum (p < 0.05). TNF-α increased 7.4–9.72-fold, whereas IL-10 rose 2.82–3.11-fold. Importantly, encapsulated antigen + alum produced the highest ELISA responses after the sixth dose (6.36-fold for MCF-7 and 7.00-fold for MDA-MB-231 versus control; p < 0.05). Hybridoma-derived antibody signals increased through day 42, and Protein G purification yielded up to ~395 μg and ~318 μg IgG. Purified antibodies reduced cell viability, and viability decreased further when antibodies were combined with doxorubicin (to ~31.6% in MCF-7 and ~40.3% in MDA-MB-231). Conclusions: Overall, sustained PCL-mediated antigen release combined with alum strengthened humoral responses to tumor lysate and enabled recovery of functional antibodies with diagnostic capture and in vitro cytotoxic activity. In future work, key mechanistic steps such as lymph-node trafficking and cross-presentation should be tested directly. Full article

Severe dengue virus (DENV) infections are associated with circulating non-neutralizing antibodies generated during heterotypic infections. Although antibodies are key mediators of both protection and pathogenesis, the specific dynamics of B cells (Bc) and their antibody responses remain insufficiently characterized due to limited methods of identifying DENV-specific Bc (DENV-Bc) and the absence of animal models resembling the human disease. Here, we developed a spectral flow cytometry assay employing biotinylated virus-like particles (VLPs) to detect DENV-Bc in C57BL/6 mice and children hospitalized with dengue. DENV-1 and DENV-2 VLPs were biotinylated, and the efficiency of biotin incorporation was assessed with an HABA-avidin assay and ELISA. Serotype specificity and optimal binding conditions were confirmed using hybridomas 4G2 (pan-flavivirus) and 3H5-1 (DENV-2 specific). Fluorescent agglutimers were subsequently generated by coupling biotinylated VLPs to streptavidin–fluorochrome complexes. Splenocytes from intraperitoneally DENV-infected mice and peripheral blood mononuclear cells (PBMCs) from naturally infected pediatric patients were stained with these VLPs and Bc-lineage markers. Biotinylated VLPs bound specifically to hybridomas, and this binding was competitively inhibited by unlabeled VLPs. After secondary DENV challenge, VLPs identified DENV-specific class-switched plasmablasts in mice. Circulating DENV-specific plasmablasts were also detected in children, with agglutimers enabling the discrimination of serotype-specific and cross-reactive responses in primary and secondary infections. This VLP-based approach represents a scalable platform to investigate the protective and pathogenic roles of DENV-Bc in infection and vaccination. Full article

Background/Objectives: The Zika virus (ZIKV) represents an ongoing threat to public health due to its neurological and congenital complications. Even after 10 years since the first major outbreak, correlated with an increase in congenital ZIKV syndrome, there is still no vaccine or treatment for this infection. Among the various existing platforms, DNA vaccines combined with the use of immunoinformatics tools allow for the efficient selection of immunogenic epitopes and immunostimulatory molecules with greater flexibility, in addition to being simple to manufacture and having a higher cost–benefit ratio in production. Methods: In this work, we conducted an integrated approach, combining in silico analyses and in vivo experimental validations, for the development of multi-epitope DNA vaccines against ZIKV. The computational analyses confirmed structural stability, adequate solubility, absence of toxicity, and immune induction potential for constructs based on epitopes from the Envelope (E) and NS1 proteins. Therefore, we evaluated DNA constructs containing the ENV + NS1 epitopes, both with and without fusion to the ssPGIP signal peptide, in BALB/c mice. Results: Both vaccines increased the population of CD4⁺ and CD8⁺ T lymphocytes, in addition to the production of IgG antibodies associated with the Th1 profile. The fusion with ssPGIP broadened the response, stimulating the release of Th1, Th2, and Th17 cytokines, as well as enhancing antibody formation. In contrast, its absence was associated with a slight increase in CD4⁺ and CD8⁺ T cells, accompanied by restricted cytokine production. Conclusions: These results indicate that epitope-targeted techniques offer a viable and safe method for inducing robust immune responses, demonstrating that combining immunoinformatics methods with early preclinical testing is an effective strategy for ZIKV vaccine development. Furthermore, although the present study focused on initial immunogenic characterization, future studies involving viral challenge in a suitable animal model will be essential to conclusively determine the protective efficacy of these vaccine candidates. Full article

Background: Venezuelan equine encephalitis virus (VEEV) poses a significant public health and biodefense threat due to periodic epidemics of severe neurological disease in the Americas, yet no licensed human vaccines or specific antiviral therapies exist. Methods: We comprehensively reviewed the current literature across three core domains: structural biology, vaccine development, and therapeutic antibodies. Results: First, we detail the complex viral structural proteome (including E1/E2 glycoproteins and Capsid), focusing on the LDLRAD3 entry receptor interaction. Second, we overview vaccine strategies, covering live-attenuated, VLP, and nucleic acid platforms designed to induce robust neutralizing antibodies. Finally, we examine the evolution of therapeutic antibodies, highlighting that optimal protection often relies on both neutralization and Fc effector functions, particularly for antibodies targeting the fusion loop or receptor-binding sites. Conclusions: Integrating structural insights with advanced antibody and vaccine engineering is essential for establishing effective clinical interventions capable of preventing future outbreaks and treating infected individuals. Full article

Background/Objectives: Notwithstanding the declaration by the World Health Organization in May 2023 regarding the conclusion of the COVID-19 pandemic, new cases of this potentially lethal infection continue to be documented globally, exerting a sustained influence on the worldwide economy and social structures. Contemporary SARS-CoV-2 variants, while associated with a reduced propensity for severe acute pathology, retain the capacity to induce long-term post-COVID syndrome, including in ambulatory patient populations. This clinical phenomenon may be attributable to potential autoimmune reactions hypothetically triggered by antiviral antibodies, thereby underscoring the need for developing novel, universal vaccines against COVID-19. The nucleocapsid protein (N), being one of its most conserved and highly immunogenic components of SARS-CoV-2, presents a promising target for such investigative efforts. However, the protective role of anti-N antibodies, generated during natural infection or through immunization with N-based vaccines, alongside the potential adverse effects associated with their production, remains to be fully elucidated. In the present study, we aim to identify potential sites of homology in structures or sequences between the SARS-CoV-2 N protein and human antigens detected using hyperimmune sera against N protein obtained from mice, rabbits, and hamsters. Methods: We employed Western blot analysis of lysates from human cell lines (MCF7, HEK293T, THP-1, CaCo2, Hep2, T98G, A549) coupled with mass spectrometric identification to assess the cross-reactivity of polyclonal and monoclonal antibodies generated against recombinant SARS-CoV-2 N protein with human self-antigens. Results: We showed that anti-N antibodies developed in mice and rabbits exhibit pronounced immunoreactivity towards specific components of the human proteome. In contrast, anti-N immunoglobulins from hamsters showed no non-specific cross-reactivity with either hamster or human proteomic extracts because of the lack of autoreactivity or immunogenicity differences. Subsequent mass spectrometric analysis of the immunoreactive bands identified principal autoantigenic targets, which were predominantly heat shock proteins (including HSP90-beta, HSP70, mitochondrial HSP60, and HSPA8), histones (H2B, H3.1–3), and key metabolic enzymes (G6PD, GP3, PKM, members of the 1st family of aldo-keto reductases). Conclusions: The results obtained herein highlight the differences in the development of anti-N humoral responses in humans and in the Syrian hamster model. These data provide a foundational basis for formulating clinical recommendations to predict possible autoimmune consequences in COVID-19 convalescents and are of critical importance for the rational design of future N protein-based, cross-protective vaccine candidates against novel coronavirus infections. Full article

Background: Children with congenital heart disease (CHD) are at substantially increased risk for respiratory infections, which occur more frequently and with greater severity than in healthy peers. This heightened vulnerability stems from multifactorial immune impairment, including defects in innate and adaptive immunity, chronic inflammation related to abnormal hemodynamics and hypoxia, reduced thymic function, and genetic syndromes affecting both cardiac and immune development. Viral pathogens—particularly respiratory syncytial virus (RSV), influenza viruses, and SARS-CoV-2—account for most infections, although bacterial pathogens remain relevant, especially in postoperative settings. Methods: This narrative review summarizes current evidence on infection susceptibility in children with CHD, the epidemiology and clinical relevance of major respiratory pathogens, and the effectiveness of available preventive measures. Literature evaluating immunological mechanisms, infection burden, vaccine effectiveness, and passive immunization strategies was examined, along with existing national and international immunization guidelines. Results: Children with CHD consistently exhibit higher rates of hospitalization, intensive care unit admission, mechanical ventilation, and mortality following respiratory infections. RSV, influenza, and SARS-CoV-2 infections are particularly severe in this population, while bacterial infections, though less common, contribute substantially to postoperative morbidity. Preventive options—including routine childhood vaccines, pneumococcal and Haemophilus influenzae type b vaccines, influenza vaccines, COVID-19 mRNA vaccines, and RSV monoclonal antibodies—demonstrate strong protective effects. New long-acting RSV monoclonal antibodies and maternal vaccination markedly enhance prevention in early infancy. However, vaccine coverage remains insufficient due to parental hesitancy, provider uncertainty, delayed immunization, and limited CHD-specific evidence. Conclusions: Respiratory infections pose a significant and preventable health burden in children with CHD. Enhancing the use of both active and passive immunization is essential to reduce morbidity and mortality. Strengthening evidence-based guidelines, improving coordination between specialists and primary care providers, integrating immunization checks into routine CHD management, and providing clear, condition-specific counseling to families can substantially improve vaccine uptake and clinical outcomes in this vulnerable population. Full article

Epizootic hemorrhagic disease (EHD) is an important livestock disease caused by Epizootic hemorrhagic disease virus (EHDV). The recent incursion and wide distribution of EHDV in Europe have increased the need for effective vaccine candidates. Background/Objectives: The VP2 protein of EHDV forms the outer capsid layer of the virion and is essential for viral assembly and host cell entry. Owing to its antigenic properties, VP2 represents a major target for vaccine development. However, the recombinant production of VP2 is limited by low stability and poor yields, representing a significant barrier for the generation of safe and effective subunit vaccines. Methods: To overcome these limitations, the VP2 protein from EHDV serotype 8 (EHDV-8) was rationally engineered with targeted modifications at both the amino and carboxyl termini of its coding sequence. Recombinant expression was performed using a baculovirus vector-mediated system in Trichoplusia ni pupae (CrisBio^® technology), employed as living biofactories. Results: The engineering of VP2 resulted in up to a tenfold increase in protein yields compared with the wild-type sequence, while maintaining the trimeric structural integrity of the recombinant protein. Both wild-type and engineered VP2 protein variants were formulated and used to immunize IFNAR(−/−) mice, a model susceptible to EHDV infection. Both engineered and wild-type VP2 formulations elicited comparable neutralizing antibody responses in vaccinated animals. Furthermore, immunization with either formulation conferred full protection against lethal EHDV-8 challenge. Conclusions: In this work, we demonstrated that the rational engineering of the VP2 protein significantly improved recombinant expression yields in a baculovirus-based system without compromising structural integrity or immunogenicity. These findings additionally demonstrate the feasibility of producing high-quality VP2 antigens in T. ni pupae using CrisBio^® technology and support their potential application in the development of subunit vaccines against EHDV. Full article

Background/Objectives: African swine fever (ASF) causes massive global swine industry losses with no effective vaccine available. This study constructed T7 phages displaying key ASFV proteins to evaluate their potential as an ASF vaccine by assessing viral shedding and immune responses in pigs. Methods: Five ASFV proteins were displayed on T7 phages to form VLPs (ASFV-SC-T7 group), with soluble proteins (ASFV-SC group) and PBS as controls; 9 piglets were immunized, boosted at 28 days, challenged with virulent ASFV, and assessed via ELISA, flow cytometry, and real-time PCR. Results: ASFV-SC-T7 induced more high-titer antibodies and elevated monocytes/CD8⁺ T cells, but all groups developed ASF lesions, with ASFV-SC-T7 having higher lung/mesenteric lymph node viral loads and no survival improvement (only delayed fever). Conclusions: T7 phage-displayed ASFV proteins activate strong immunity, confirming T7 phages as a viable delivery platform, but failed to protect against virulent ASFV, requiring future optimization of antigens and regimens. Full article

RNA-based vaccination has been broadly applied in the COVID-19 pandemic. A characteristic of the immunization was fast-waning immunity. However, the time scale of this process varied considerably for virus subtypes and among individuals. Understanding the origin of this variability is crucial in order to improve future vaccination strategies. Here, we introduce a mathematical model of RNA-based vaccination and the kinetics of the induced immune response. In the model, antigens produced following vaccination give rise to an immune response leading to germinal center reactions and accordingly B-cell differentiation into memory B-cells and plasma cells. In a negative feedback loop, the antibodies synthesized by newly specified plasma cells shut down the germinal center reaction as well as antigen-induced differentiation of memory B-cell into plasma cells. This limits the build-up of long-lasting immunity and thus is accompanied by fast-waning immunity. The detailed data available on infection with and vaccination against SARS-CoV-2 enabled computational simulation of essential processes of the immune response. Through simulation, we analyzed to what extent a single- or double-dose vaccination provides protection against infection. We find that variability in the immune response in individuals, originating, e.g., in different immune-cell densities, results in a broad log-normal-like distribution of the vaccine-induced protection times that peaks around 100 days. Protection times decrease for virus variants with mutated antibody-binding sites or increased replication rates. Independent of these virus specifics, our simulations suggest optimal timing of a second dose about 5 weeks after the first in agreement with clinical trials. Full article